MA and gp120 induced the expression of CYP
As CYP has been shown to be involved in oxidative stress in many organs/tissues including the brain, we measured the mRNA expression levels of various CYPs in astrocytes treated with MA and/or gp120. Both MA and gp120 induced different isozymes of CYP at variable levels (Figure 4a). Among the isozymes induced, CYP2E1 (1.7±0.2-fold), CYP2D6 (2.3±0.3-fold), and CYP2B6 (3.2±03-fold) clearly showed additive increases (two-way ANOVA showed P-value=0.34, 0.18, and 0.84, respectively, suggesting the absence of synergy) in the levels of mRNA expression with gp120+MA when compared with either MA (nonsignificant change for CYP2E1 and CYP2D6 and 2.8±0.3-fold for CYP2B6) or gp120 (1.3±0.1-fold for CYP2E1 and 1.8±0.3-fold for CYP2D6 and nonsignificant for CYP2B6) alone (Figure 4a). These results were further confirmed at the protein level for CYP2E1 (1.3±0.1-fold for gp120+MA), CYP2D6 (1.7±0.2-fold for gp120+MA), and CYP2B6 (1.4±0.1-fold for gp120+MA) (Figure 4b). Furthermore, to confirm this phenomenon in primary astrocytes, human fetal astrocytes (HFA) were treated with gp120 IIIB and/or MA and the protein and mRNA expression levels of the CYPs were measured. Similar to SVGA astrocytes, gp120 and MA also showed additive increase in the levels of CYP2E1, CYP2B6, and CYP2D6 mRNA (Figure 4c) and protein (Figure 4d) in HFA primary cells. The analysis run for synergy using two-way ANOVA showed nonsignificant P-values, suggesting no synergistic interaction.